|
| Status |
Public on Jan 31, 2020 |
| Title |
SLX.14720.A009 |
| Sample type |
SRA |
| |
|
| Source name |
Drosophila Melanogaster Elav x PrP (Cyt) whole head homogenate treated with Normal Brain Homogenate, harvested at 5 days of age
|
| Organism |
Drosophila melanogaster |
| Characteristics |
tissue: Whole head homogenate
fly line: Elav x PrP (Cyt)
age in days: 5
treatment: Normal Brain Homogenate
|
| Treatment protocol |
Drosophila at were Drosophila at the larval stage of development were exposed to brain homogenates of cerebral cortex tissue from confirmed scrapie-positive or known scrapie-negative sheep. After hatching, Drosophila were transferred to inoculum-free tubes and allowed to develop normally. At 5 days and 30/40 days post hatching, the flies were euthanized and then decapitated for RNA extraction
|
| Extracted molecule |
total RNA |
| Extraction protocol |
15 fly heads was homogenised by manual grinding in an eppendorf tube with 50μl Trizol (Cat No. 15596-018; Invitrogen). A further 50μl of Trizol were mixed with the homogenate, which was then microfuged at 16,160 x g for 10 minutes and the supernatant transferred to RNAse-free tubes before adding a further 50μl Trizol and incubated for 5 minutes. RNA was isolated by conventional chloroform:isoamylalcohol:isopropanol extraction followed by ethanol precipitation. Air-dried RNA pellets were treated with DNAse 1 (Cat No. EN0521; Thermo Scientific) at 37 ̊C for 60 minutes, with the reaction halted by the addition of EDTA and incubation at 65 ̊C for 10 minutes. RNA samples were then subjected to the Qiagen RNeasy RNA Cleanup protocol according to the manufacturers’ instructions. The RNA samples were eventually eluted from RNeasy mini-spin columns, ethanol precipitated, re-suspended in RNAse-free water and stored at -80 ̊C.
A library of mRNA was prepared from 200 ng of total RNA, extracted as described above and that had been originally pooled from 15 fly heads from each experimental condition, using the Illumina TruSeq Stranded mRNA Library Prep kit (Illumina, San Diego, CA, USA) according to the manufacturer’s protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
RNA from Drosophila Melanogaster Elav x PrP (Cyt) whole head homogenate treated with Normal Brain Homogenate, harvested at 5 days of age
|
| Data processing |
Fastq reads were mapped to Drosophila Genome (built BDGP6) using Tophat V2.0.11
htseq-count 0.6.1p1 was used to generate gene level counts based on Ensembl V77 GTF
Differential gene expression was performed using edgeR
Genome_build: BDGP6
Supplementary_files_format_and_content: tab-delimited text containing raw gene-level count for each sample
|
| |
|
| Submission date |
Jan 21, 2020 |
| Last update date |
Jan 31, 2020 |
| Contact name |
Brian Yee Hong Lam |
| E-mail(s) |
yhbl2@cam.ac.uk
|
| Organization name |
University of Cambridge
|
| Department |
Metabolic Research Laboratories
|
| Street address |
Box 289 Addenbookes Hospital, Hills Road
|
| City |
Cambridge |
| ZIP/Postal code |
CB2 0QQ |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL21306 |
| Series (1) |
| GSE144028 |
Transcriptional signature of prion-induced neurotoxicity in a Drosophila model of transmissible mammalian prion disease |
|
| Relations |
| BioSample |
SAMN13898240 |
| SRA |
SRX7587186 |
